Project Title: An Orbitrap Mass Spectrometry System for the University of Cincinnati Proteomics Laboratory Project Summary/Abstract This proposal requests funding for a nanoflow LC system coupled to a Q Exactive HF-X mass spectrometry system to be housed and used in the University of Cincinnati Proteomics Laboratory (UCPL). This system will support the NIH-funded research community across UC, Cincinnati Children's Hospital Medical Center (CCHMC) and the Cincinnati VA Medical Center. The new instrument is requested to maintain the high level of mass spectrometry-based proteomics support for the research community, which encompasses over 300 NIH- funded investigators with over 200 million dollars of support annually. The new instrument is needed to both improve the technical capabilities through increased sensitivity and to lessen the extended wait times for instrument access due to the increased number and complexity of research projects supported by the UCPL. The research support provided by the UCPL has expanded well beyond routine sample analysis and is now most often directed at more complex, research-oriented or sample-intensive studies that require the best sensitivity and greater instrument time. To support these projects, the UCPL is currently equipped with two mass spectrometers: 1) Sciex 4800 MALDI-TOF/TOF MS and 2) Sciex 5600+ Q-Tof, nanoLC-MS/MS system. However, the demand for nanoLC-MS/MS instrument has resulted in sample queues that are often 2-3 weeks long, which dramatically affects the ability to support the 30+ investigators each year. In addition, the MALDI- TOF/TOF system (purchased in 2005) has progressed to the point where the vendor can no longer ensure that the system can be repaired, and thus many of the routine analyses currently done on this system are being migrated to the only other available system, adding to the wait times. Specific areas of research for the Major User Groups include, global profiling of regulatory modifications such as phosphorylation and ubiquitination, detailed structural characterization of lipoproteins through cross-linking studies, evaluation and quantification of differential proteins in brain synapse function, and evaluation of large patient cohorts for the identification, quantification and validation of biomarkers of kidney diseases. The PI of the application has a long history (20+ years) of success in developing, adapting and applying biological mass spectrometry to solve research questions. More importantly, he works effectively at the interface of technology and biology as a collaborative partner with investigators to design experiments to maximize success. The requested system will be operated by a PhD scientist with 11 years of experience in all aspects of nanoLC-MS/MS analysis. Importantly, the UCPL is supported institutionally by the University of Cincinnati to offset the cost of service contracts and technology development and to help minimize the direct cost of services to the investigators. The requested instrument is ideally suited to address both the technical specifications and capacity issues for the 4 Major User Groups encompassing 8 NIH funded investigators.